The goal of this proposal is to identify the genes responsible for cholesterol gallstone formation and their function. Identification of these genes in the inbred mouse may define the pathophysiological mechanisms contributing to disease and may identify key proteins as targets for rational drug design. Aim 1 proposes to use transgenic or knockout technology to prove the identity of three strong candidate genes discovered during the last proposal; Abcb11 for Lith1, Abcc2 for Lith2, and carboxypeptidase E (Cpe), a mutation that results in both obesity and increased gallstone susceptibility. Aim 2 proposes to test promising candidate genes in the remaining QTL regions discovered during the last grant period by comparing the gallstone-resistant and susceptible strains for sequence and expression differences. To find additional genes associated with susceptibility to gallstone formation, aim 3 proposes to carry out a series of crosses between gallstone susceptible and resistant strains. Analysis will include both main QTL and interacting or epistatic QTL using our recently developed statistical methodology. Identifying the full range of QTLs that lead to gallstone disease in the mouse will position the field of cholesterol gallstone research to take advantage of the large genomic sequencing efforts currently underway.